Non-invasive pre-natal screening (NIPT) by sequencing of cell-free fetal DNA (cffDNA) isolated from maternal blood has resulted in significant benefits; the screening is safe, relatively low cost, and greatly reduces the number of invasive amniocentesis and chorionic villus sampling (CVS) procedures. Current NIPT screens have a very high negative predictive value (>99%) but have a relatively low positive predictive value; there is a high likelihood (>80% in some cases) of a false positive result. Therefore, a follow-up invasive test is required to confirm positive NIPT screens. Furthermore, low sensitivity limits NIPT screening to trisomies that are more easily measured from the low levels of circulating fetal DNA in maternal blood. A test that can simultaneously detect trisomies as well as subchromosomal abnormalities, without the need for follow-up testing, would represent a significant advance of the current standard of care. This project proposes to develop a cell-based NIPT test that provides a true diagnosis of trisomies without the need for follow-up amniocentesis or CVS. Furthermore, the test would have sufficient gains in sensitivity and specificity to allow detection of rare subchromosomal genetic disorders, resulting in a significant advance in terms of the current non-invasive screening capabilities. The workflow of the test takes advantage of several technologies that we have pioneered: rare cell isolation, single-cell manipulation/analysis, and single-cell sequencing. Initial proof-of-concept experiments have demonstrated that model cells representing fetal trophoblasts can be isolated and detected, and we have demonstrated the ability to isolate and analyze individual cells via NGS from an enriched, heterogeneous mixture. The proposed workflow comprises a cell isolation system, library prep kits for sequencing single cells on commercially available sequencers, and bioinformatics software for analysis. Successful execution of this project will result in a complete workflow for genetic analysis of cffDNA samples, resulting in a true in vitro diagnostic test that can detect not only trisomies but other rare subchromosomal genetic disorders, without requiring an invasive follow-up analysis. The development of this capability will represent a substantial advance in the current standard of care and will also serve as a platform for development of another rare-cell testing.